The long-term objective is to understand synaptic mechanisms in the central nervous system, particularly in instances where there might be multiple receptors for a transmitter or the possibility for interactions between transmitters, such as might occur at polyinnervated neurons. This proposal is specifically concerned with properties of glycine receptors and glycine-activated chloride channels ln cultured embryonic rat modullary neurons. The basic experimental design involves voltage-clamping the cultured cells using the whole-cell gigaseal technique. Agonist doseresponse relationships under different experimental conditions will be obtained by using pressure ejection from a second pipette or bath application. Specific Aim I is to characterize the multiple populations of glycine receptors. Questions to be asked include: a) can these receptors be further distinguished on the basis of responses evoted by different aqonists and their strychnine sensitivities, and b) do both receptor classes co-exist on single cells? These questions will be addressed by obtaining agonist dose-response curves in the presence of different strychnine concentrations. Also, stationary fluctuation analysis will be used to provide a macroscopic description of the channels activated via the receptor subtypes. Specific Aim II is to study, quantitatively, interactions between glycine and its various possible agonists. Dose-response relationships of one agonist, in the presence of a constant concentration of another ligand, will be determined. It may be possible to find aqonists specific for the glycine receptor subtypes. Specific Aim II is to characterize the channels activated in rat medullary neurons by glycine and other agonists. Both stationary fluctuation analysis and single channel recordings will be used to determine characteristics of chloride channels activated by the two different receptor subtypes. Parameters to be measured include the dominant and average single channel conductance and mean channel lifetime. Open and closed time distributions and amplitude histograms will also be generated. Since certain experimental disorders, e.g. spasticities, neuropathologies are associated with disturbances in glycinergic mechanisms, a detailed study of supraspinal glycine receptors and the channels they gate may provide information useful in treating related neuropathologies in humans.